Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

The Rad50 zinc-hook is a structure joining Mre11 complexes in DNA recombination and repair

Nature volume 418, pages 562–566 (2002)Cite this article

Abstract

The Mre11 complex (Mre11–Rad50–Nbs1) is central to chromosomal maintenance and functions in homologous recombination, telomere maintenance and sister chromatid association1,2,3,4,5,6,7. These functions all imply that the linked binding of two DNA substrates occurs, although the molecular basis for this process remains unknown. Here we present a 2.2 Å crystal structure of the Rad50 coiled-coil region that reveals an unexpected dimer interface at the apex of the coiled coils in which pairs of conserved Cys-X-X-Cys motifs form interlocking hooks that bind one Zn2+ ion. Biochemical, X-ray and electron microscopy data indicate that these hooks can join oppositely protruding Rad50 coiled-coil domains to form a flexible bridge of up to 1,200 Å. This suggests a function for the long insertion in the Rad50 ABC-ATPase domain8. The Rad50 hook is functional, because mutations in this motif confer radiation sensitivity in yeast and disrupt binding at the distant Mre11 nuclease interface. These data support an architectural role for the Rad50 coiled coils in forming metal-mediated bridging complexes between two DNA-binding heads. The resulting assemblies have appropriate lengths and conformational properties to link sister chromatids in homologous recombination and DNA ends in non-homologous end-joining.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on SpringerLink
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Rad50 domains, sequence conservation of the CXXC motif and Zn2+-mediated dimerization of Rad50.
Figure 2: Crystal structures of the central portion of the Rad50 coiled coil reveal a dimerization motif mediated by a single Zn2+-binding site.
Figure 3: Negatively stained electron micrographs of human and P. furiosus Mre11 complexes reveal conformational properties of the component domains including Zn2+-mediated tail-to-tail linkages between individual coiled-coil arms of Rad50.
Figure 4: In vivo effects of cysteine mutations in the CXXC motif and biological implications for metal-mediated dimerization of Rad50.

Similar content being viewed by others

References

  1. Haber, J. E. The many interfaces of Mre11. Cell 95, 583–586 (1998)

    Article  CAS  Google Scholar 

  2. Bressan, D. A., Baxter, B. K. & Petrini, J. H. The Mre11–Rad50–Xrs2 protein complex facilitates homologous recombination-based double-strand break repair in Saccharomyces cerevisiae. Mol. Cell. Biol. 19, 7681–7687 (1999)

    Article  CAS  Google Scholar 

  3. Moreau, S., Ferguson, J. R. & Symington, L. S. The nuclease activity of Mre11 is required for meiosis but not for mating type switching, end joining, or telomere maintenance. Mol. Cell. Biol. 19, 556–566 (1999)

    Article  CAS  Google Scholar 

  4. Yamaguchi-Iwai, Y. et al. Mre11 is essential for the maintenance of chromosomal DNA in vertebrate cells. EMBO J. 18, 6619–6629 (1999)

    Article  CAS  Google Scholar 

  5. Tsukamoto, Y., Taggart, A. K. & Zakian, V. A. The role of the Mre11–Rad50–Xrs2 complex in telomerase-mediated lengthening of Saccharomyces cerevisiae telomeres. Curr. Biol. 11, 1328–1335 (2001)

    Article  CAS  Google Scholar 

  6. Merino, S. T., Cummings, W. J., Acharya, S. N. & Zolan, M. E. Replication-dependent early meiotic requirement for Spo11 and Rad50. Proc. Natl Acad. Sci. USA 97, 10477–10482 (2000)

    Article  ADS  CAS  Google Scholar 

  7. Hartsuiker, E., Vaessen, E., Carr, A. M. & Kohli, J. Fission yeast Rad50 stimulates sister chromatid recombination and links cohesion with repair. EMBO J. 20, 6660–6671 (2001)

    Article  CAS  Google Scholar 

  8. Hopfner, K. P. et al. Structural biochemistry and interaction architecture of the DNA double-strand break repair Mre11 nuclease and Rad50-ATPase. Cell 105, 473–485 (2001)

    Article  CAS  Google Scholar 

  9. Petrini, J. H. The Mre11 complex and ATM: collaborating to navigate S phase. Curr. Opin. Cell Biol. 12, 293–296 (2000)

    Article  CAS  Google Scholar 

  10. Huang, J. & Dynan, W. S. Reconstitution of the mammalian DNA double-strand break end-joining reaction reveals a requirement for an Mre11/Rad50/NBS1-containing fraction. Nucleic Acids Res. 30, 667–674 (2002)

    Article  CAS  Google Scholar 

  11. Furuse, M. et al. Distinct roles of two separable in vitro activities of yeast Mre11 in mitotic and meiotic recombination. Embo J 17, 6412–6425 (1998)

    Article  CAS  Google Scholar 

  12. Paull, T. T. & Gellert, M. The 3′ to 5′ exonuclease activity of Mre 11 facilitates repair of DNA double-strand breaks. Mol. Cell 1, 969–979 (1998)

    Article  CAS  Google Scholar 

  13. Connelly, J. C., de Leau, E. S. & Leach, D. R. DNA cleavage and degradation by the SbcCD protein complex from Escherichia coli. Nucleic Acids Res. 27, 1039–1046 (1999)

    Article  CAS  Google Scholar 

  14. Hopfner, K. P. et al. Mre11 and Rad50 from Pyrococcus furiosus: cloning and biochemical characterization reveal an evolutionarily conserved multiprotein machine. J. Bacteriol. 182, 6036–6041 (2000)

    Article  CAS  Google Scholar 

  15. Trujillo, K. M. & Sung, P. DNA structure-specific nuclease activities in the Saccharomyces cerevisiae Rad50/Mre11 complex. J. Biol. Chem. 276, 35458–35464 (2001)

    Article  CAS  Google Scholar 

  16. Lobachev, K. S., Gordenin, D. A. & Resnick, M. A. The Mre11 complex is required for repair of hairpin-capped double-strand breaks and prevention of chromosome rearrangements. Cell 108, 183–193 (2002)

    Article  CAS  Google Scholar 

  17. Xiao, Y. & Weaver, D. T. Conditional gene targeted deletion by Cre recombinase demonstrates the requirement for the double-strand break repair Mre11 protein in murine embryonic stem cells. Nucleic Acids Res. 25, 2985–2991 (1997)

    Article  CAS  Google Scholar 

  18. Luo, G. et al. Disruption of mRad50 causes embryonic stem cell lethality, abnormal embryonic development, and sensitivity to ionizing radiation. Proc. Natl Acad. Sci. USA 96, 7376–7381 (1999)

    Article  ADS  CAS  Google Scholar 

  19. Zhu, J., Petersen, S., Tessarollo, L. & Nussenzweig, A. Targeted disruption of the Nijmegen breakage syndrome gene NBS1 leads to early embryonic lethality in mice. Curr. Biol. 11, 105–109 (2001)

    Article  CAS  Google Scholar 

  20. de Jager, M. et al. Human Rad50/Mre11 is a flexible complex that can tether DNA ends. Mol. Cell 8, 1129–1135 (2001)

    Article  CAS  Google Scholar 

  21. Chen, L., Trujillo, K., Ramos, W., Sung, P. & Tomkinson, A. E. Promotion of Dnl4-catalyzed DNA end-joining by the Rad50/Mre11/Xrs2 and Hdf1/Hdf2 complexes. Mol. Cell 8, 1105–1115 (2001)

    Article  CAS  Google Scholar 

  22. Hopfner, K. P. et al. Structural biology of Rad50 ATPase: ATP-driven conformational control in DNA double-strand break repair and the ABC-ATPase superfamily. Cell 101, 789–800 (2000)

    Article  CAS  Google Scholar 

  23. Connelly, J. C., Kirkham, L. A. & Leach, D. R. The SbcCD nuclease of Escherichia coli is a structural maintenance of chromosomes (SMC) family protein that cleaves hairpin DNA. Proc. Natl Acad. Sci. USA 95, 7969–7974 (1998)

    Article  ADS  CAS  Google Scholar 

  24. Anderson, D. E., Trujillo, K. M., Sung, P. & Erickson, H. P. Structure of the Rad50 × Mre11 DNA repair complex from Saccharomyces cerevisiae by electron microscopy. J. Biol. Chem. 276, 37027–37033 (2001)

    Article  CAS  Google Scholar 

  25. Sharples, G. J. & Leach, D. R. Structural and functional similarities between the SbcCD proteins of Escherichia coli and the RAD50 and MRE11 (RAD32) recombination and repair proteins of yeast. Mol. Microbiol. 17, 1215–1217 (1995)

    Article  CAS  Google Scholar 

  26. Hirano, T. SMC-mediated chromosome mechanics: a conserved scheme from bacteria to vertebrates? Genes Dev. 13, 11–19 (1999)

    Article  CAS  Google Scholar 

  27. Wu, X. & McMurray, C. T. Calmodulin kinase II attenuation of gene transcription by preventing cAMP response element-binding protein (CREB) dimerization and binding of the CREB-binding protein. J. Biol. Chem. 276, 1735–1741 (2001)

    Article  CAS  Google Scholar 

  28. Brunger, A. T. et al. Crystallography & NMR system: A new software suite for macromolecular structure determination. Acta Crystallogr. D 54, 905–921 (1998)

    Article  CAS  Google Scholar 

  29. Bressan, D. A., Olivares, H. A., Nelms, B. E. & Petrini, J. H. Alteration of N-terminal phosphoesterase signature motifs inactivates Saccharomyces cerevisiae Mre11. Genetics 150, 591–600 (1998)

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Usui, T., Ogawa, H. & Petrini, J. H. A DNA damage response pathway controlled by Tel1 and the Mre11 complex. Mol. Cell 7, 1255–1266 (2001)

    Article  CAS  Google Scholar 

Download references

Acknowledgements

We thank D. Turk for MAIN; M. Gehl for atomic absorption spectroscopy analysis and A.-M. Hayes for technical advice. We acknowledge the key facilities and staff of synchrotron beamlines BL9-2 (SSRL) and 5.0.2 (ALS). Key funding was provided by the US Department of Energy (DOE) and NCI (J.A.T. and J.P.C), the American Cancer Society (J.P.C.), the Canadian Institutes of Health Research (L.C.), National Institutes of Health, Human Frontiers Science Program and DOE (J.H.J.P), Skaggs Institute for Chemical Biology (K.P.H.), the National Institutes of Mental Health (C.T.M.) and The Swiss National Science Foundation (J.L.B).

Author information

Author notes

  1. Karl-Peter Hopfner and Lisa Craig: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Molecular Biology and Skaggs Institute for Chemical Biology, The Scripps Research Institute, La Jolla, California, 92037, USA

    Lisa Craig, Gabriel Moncalian, Jean-Luc Bodmer & John A. Tainer

  2. Gene Center and Institute of Biochemistry, University of Munich, 81377, Munich, Germany

    Karl-Peter Hopfner & Annette Karcher

  3. Laboratory of Genetics, University of Wisconsin, Madison, Wisconsin, 53706, USA

    Robert A. Zinkel

  4. Molecular Biology, Memorial Sloan-Kettering Cancer Center, New York, New York, 10021, USA

    Takehiko Usui & John H. J. Petrini

  5. Department of Molecular Pharmacology and Experimental Therapeutics, Mayo Clinic and Foundation, Rochester, Minnesota, 55905, USA

    Barbara A. L. Owen & Cynthia T. McMurray

  6. The Radiation Oncology Research Laboratory, University of Maryland School of Medicine, Baltimore, Maryland, 21201, USA

    Brendan Henderson & James P. Carney

  7. Life Sciences Division, Lawrence Berkeley National Laboratory, Berkeley, California, 94720, USA

    John A. Tainer

Authors
  1. Karl-Peter Hopfner
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Lisa Craig
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Gabriel Moncalian
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Robert A. Zinkel
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Takehiko Usui
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Barbara A. L. Owen
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Annette Karcher
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Brendan Henderson
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Jean-Luc Bodmer
    View author publications

    You can also search for this author in PubMed Google Scholar

  10. Cynthia T. McMurray
    View author publications

    You can also search for this author in PubMed Google Scholar

  11. James P. Carney
    View author publications

    You can also search for this author in PubMed Google Scholar

  12. John H. J. Petrini
    View author publications

    You can also search for this author in PubMed Google Scholar

  13. John A. Tainer
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to John H. J. Petrini or John A. Tainer.

Ethics declarations

Competing interests

The authors declare that they have no competing financial interests.

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Cite this article

Hopfner, KP., Craig, L., Moncalian, G. et al. The Rad50 zinc-hook is a structure joining Mre11 complexes in DNA recombination and repair. Nature 418, 562–566 (2002). https://doi.org/10.1038/nature00922

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nature00922

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing